Determination of Intrinsic Creatine Transporter (Slc6a8) Activity and Creatine Transport Function of Leukocytes in Rats.

Creatine transporter (CRT) deficiency (CRT-D) results in a significant reduction of brain creatine levels, which causes various neurological symptoms in early childhood, and diagnosis of the severity of CRT-D based on the residual CRT transport activity in liquid biopsy samples would be beneficial for early intervention. The apparent reduction in creatine transport activity in CRT-D is thought to be due to reduced intrinsic CRT-mediated creatine transport per CRT protein and/or reduced absolute CRT protein expression on the plasma membranes. The purpose of this study was thus to determine the normal level of intrinsic CRT-mediated creatine transport activity based on absolute CRT protein quantification using rat CRT-overexpressing HEK293 cells (CRT/HEK293 cells), and to clarify creatine transport in erythrocyte- and leukocyte-enriched fractions isolated from the circulating blood of rats.

Developmental changes in transporter and receptor protein expression levels at the rat blood-brain barrier based on quantitative targeted absolute proteomics.

The blood-brain barrier (BBB) transport systems regulate the supply of nutrients, amino acids, vitamins, and hormones to the developing brain, as well as blocking the entry of xenobiotics and drugs. The purpose of this study was to clarify the developmental changes in the absolute protein expression levels of BBB transport-related proteins in developing rat brain capillaries, using quantitative targeted absolute proteomics (QTAP). The changing patterns of ATP-binding cassette (ABC) and solute carrier (SLC) transporters, receptors, and tight junction/adherence junction-related proteins were classified into 4 types: uphill (continuously increasing expression from postnatal day (P) 1 to P56), bell-shape/inverted bell-shape (increased/decreased expression from P1 to P14 followed by decreased/increased expression from P21 to P56), downhill (continuously decreasing expression from P1 to P56), and constant (no significant difference from P1 to P56).

Distinct Transport Properties of Human Pannexin 1 and Connexin 32 Hemichannels.

Pannexin (Px) and connexin (Cx) hemichannels mediate bidirectional membrane transport in response to various stimuli and are involved in drug efficacy and toxicity. The purpose of the present study was to clarify in detail the transport characteristics of Px1 and Cx32 hemichannels by establishing transport assay systems using human Px1- and P2RX7 receptor-overexpressing HEK293 cells (Px1/P2RX7/HEK293) and Cx32-overexpressing HEK293 cells (Cx32/HEK293), in which P2RX7 and an extracellular Ca-depleted condition serve as the opening trigger, respectively. Uptake of the cationic fluorescent dye propidium iodide (PI) was significantly increased in Px1/P2RX7/HEK293 cells compared to that in mock cells, whereas there was no significant uptake of the anionic fluorescent dye sulforhodamine 101 (SR101).